Function of novel gene, EAT that is early gene induced by all-trans retinoic acid in human EC cells

本研究的目的是分离维甲酸（RA）诱导人EC细胞NCR-G3细胞分化的必需基因。我们分离到一个新的基因，EAT，它在分化早期短暂表达，与细胞死亡预防分子bcl-2具有同源性。通过分析这个新基因EAT，我们得到了以下结果。为了阐明EAT在正常和胎儿组织中的表达，利用重组蛋白生成了小鼠单克隆抗体3A2。通过免疫组织化学研究，在多种人类胎儿和成人组织中观察到EAT的表达，包括上皮细胞、造血细胞和内分泌细胞。在小鼠胚胎中，在受精后快速诱导，在2 - 8细胞阶段达到最高水平，然后在囊胚中降低到未受精卵水平以下。2)我们确定了是否像Bcl-2报道的那样，EAT可以抑制化疗药物诱导的凋亡细胞死亡。转染了EAT的细胞对CDDP或其他化疗药物有更高的耐药性。用CDDP和卡铂治疗后，亲代细胞的DNA断裂呈浓度依赖性。相比之下，用同样浓度的这些药物治疗后，转染了EAT的细胞没有出现DNA断裂。在EAT转基因小鼠中未发生CDDP诱导的急性肾小管坏死。这些结果表明，EAT可能通过其抗凋亡功能在早期胚胎发生和细胞序贯功能中发挥重要作用。

The aim of this study was to isolate the essential genes in retinoic acid (RA)-induced differentiation of human EC cells, NCR-G3 cell. We isolated a novel gene, EAT which transiently expressed in the early stage of differentiation and has homology with bcl-2, a cell death-preventing molecule. We obtained following results by analyzing this novel gene, EAT.To elucidate expression of EAT in normal and fetal tissue, a murine monoclonal antibody, 3A2 was generated by use of recombinant protein to EAT. By immnohistochemical studies, EAT expression was observed in a wide variety of human fetal and adult tissues, including those of epithelial, hematopoietic and endocrine cells. In murine embryos, EAT was induced rapidly after fertilization reaching maximum level at the from the 2 - to 8-cell stage and then decrease to below unfertilized egg levels in blastocyst.2) We determined whether the apoptotic cell death induced by chemotherapeutic agents could be inhibited by EAT as is reported with Bcl-2. Cells transfected with EAT showed higher resistance to CDDP or among other chemotherapeutic agents tested. DNA fragmentation of the parental cells following treatment with CDDP and carboplatin was observed in a concentration-dependent manner. In contrast, cells transfected with EAT did not show DNA fragmentation following treatment with the same concentration of these drugs. Acute tubular necrosis induced by CDDP administration did not occur in the EAT transgenic mice.These results suggest that EAT may play an important role in early embryogenesis and sequential cell function by its anti-apoptotic functions.